Anti-axl antibodies, antibody fragments and their immunoconjugates and uses thereof

Abstract

An antibody or antibody fragment having a heavy chain variable region and/or light chain variable region that specifically binds to Ax1 protein. Immunoconjugates, pharmaceutical compositions and kits comprising the antibodies and antibody fragments are also provided. Also disclosed are methods of treating Ax1-expressing cancers using the antibodies, antibody fragments, immunoconjugates and pharmaceutical compositions of the present invention.

1 .- 20 . (canceled) 
     
     
         21 . A method of treating an AXL protein expressing cancer comprising administering to a patient a therapeutically effective amount of an immunoconjugate comprising
 an antibody or antibody fragment that specifically binds to Ax1 protein, said antibody or antibody fragment comprising a heavy chain variable region including three complementarity determining regions having H1, H2, and H3 sequences, and a light chain variable region including three complementarity determining regions having L1, L2, and L3 sequences, wherein:   (a) the H1 sequence is X 1 GX 2 X 3 MX 4  (SEQ ID NO: 1);   (b) the H2 sequence is LIKX 5 SNGGTX 6 YNQKFKG (SEQ ID NO: 2); and   (c) the H3 sequence is GX 7 X 8 X 9 X 10 X 11 X 12 X 13 X 14 DYX 15 X 16  (SEQ ID NO: 3),   
       wherein
 X 1  is T or A or W, 
 X 2  is H or A, 
 X 3  is T or I, 
 X 4  is N or I, 
 X 5  is P or N, 
 X 6  is S or I or T, 
 X 7  is H or D or E or P or R or W, 
 X 8  is Y or N, 
 X 9  is E or A or D or F or G or H or I or L or M or N or R or V or Y, 
 X 10  is S or D or M or N or Q, 
 X 11  is Y or C or E or P, 
 X 12  is F or E or N or S or T or V, 
 X 13  is A or D or G or L or Y, 
 X 14  is M or E or F, 
 X 15  is W or A or D or H or L or N or P or R or T, 
 X 16  is G or H, and 
 (d) the L1 sequence is KASQDX 17 X 18 SX 19 VX 20  (SEQ ID NO: 4); 
 (e) the L2 sequence is X 21 X 22 X 23 TRX 24 T (SEQ ID NO: 5); and 
 (f) the L3 sequence is QEX 25 X 26 SX 27 X 28 X 29 X 30  (SEQ ID NO: 6), 
 
       wherein
 X 17  is V or D or G or N or W, 
 X 18  is S or V, 
 X 19  is A or L or M, 
 X 20  is A or D or N or Q, 
 X 21  is W or F, 
 X 22  is A or I or N or P or Q, 
 X 23  is S or D, 
 X 24  is H or D, 
 X 25  is H or C or F or I or L or Q or S or T or V or Y, 
 X 26  is F or C or D or E or G or N or S, 
 X 27  is T or C or P, 
 X 28  is P or A or C or D or E or H or K or S or T or V or W, 
 X 29  is L or G or R, and 
 X 30  is T or I or R; and 
 at least one agent selected from the group consisting of a chemotherapeutic agent, a radioactive atom, a cytostatic agent and a cytotoxic agent; and 
 wherein the AXL protein expressing cancer is selected from the group consisting of small cell lung cancer, non-small cell lung cancer, renal cancer, liver cancer, melanoma, bladder cancer, cervical cancer, hepatic carcinoma, gastric cancer, sarcoma and kidney cancer. 
 
     
     
         22 . The method of  claim 21 , wherein the heavy chain variable region of the antibody or antibody fragment is encoded by a DNA sequence selected from the group consisting of sequences of SEQ ID NOS: 11-13. 
     
     
         23 . The method of  claim 21 , wherein the light chain variable region of the antibody or antibody fragment is encoded by a DNA sequence selected from the group consisting of sequences of SEQ ID NOS: 7-10. 
     
     
         24 . The method of  claim 21 , wherein the heavy chain variable region of the antibody or antibody fragment is encoded by the DNA sequence of SEQ ID NO: 12 and the light chain variable region of the antibody or antibody fragment is encoded by the DNA sequence of SEQ ID NO: 9. 
     
     
         25 . The method of  claim 21 , wherein the antibody or antibody fragment has a higher binding affinity to the Ax1 protein at a value of a condition in a tumor microenvironment in comparison with a different value of the same condition that occurs in a non-tumor microenvironment. 
     
     
         26 . The method of  claim 25 , wherein the condition is pH. 
     
     
         27 . The method of  claim 26 , wherein the pH in the tumor microenvironment is in a range of from 5.8 to 7.0 and the pH in the non-tumor microenvironment is in a range of 7.2-7.6. 
     
     
         28 . The method of  claim 21 , wherein the antibody or antibody fragment has a ratio of binding affinity to the Ax1 protein at a value of a condition in a tumor microenvironment to a binding affinity to the Ax1 protein at a different value of the same condition in a non-tumor microenvironment of at least about 1.5:1. 
     
     
         29 . The method of  claim 21 , wherein the antibody or antibody fragment is selected from the group consisting of a chimeric antibody, a multispecific antibody and a humanized antibody. 
     
     
         30 . The method of  claim 21 , wherein the immunoconjugate comprises at least two agents. 
     
     
         31 . The method of  claim 21 , wherein the antibody or antibody fragment and the at least one agent are covalently bonded to a linker molecule. 
     
     
         32 . The method of  claim 21 , wherein the at least one agent is selected from the group consisting of maytansinoids, auristatins, dolastatins, calicheamicin, pyrrolobenzodiazepines, and anthracyclines. 
     
     
         33 . The method of  claim 21 , wherein the cancer is non-small cell lung cancer. 
     
     
         34 . The method of  claim 21 , wherein the cancer is sarcoma. 
     
     
         35 . The method of  claim 34 , wherein the sarcoma is osteosarcoma. 
     
     
         36 . The method of  claim 22 , wherein the light chain variable region of the antibody or antibody fragment is encoded by a DNA sequence selected from the group consisting of sequences of SEQ ID NOS: 7-10. 
     
     
         37 . The method of  claim 27 , wherein the pH in the tumor microenvironment is about 6.0 and the pH in the non-tumor microenvironment is about 7.4. 
     
     
         38 . The method of  claim 22 , wherein the antibody or antibody fragment has a ratio of binding affinity to the Ax1 protein at a pH of 5.8 to 7.0 of a tumor microenvironment to a binding affinity to the Ax1 protein at a pH of 7.2 to 7.6 in a non-tumor microenvironment of at least about 1.5:1. 
     
     
         39 . The method of  claim 36 , wherein the antibody or antibody fragment has a ratio of binding affinity to the Ax1 protein at a pH of 5.8 to 7.0 of a tumor microenvironment to a binding affinity to the Ax1 protein at a pH of 7.2 to 7.6 in a non-tumor microenvironment of at least about 1.5:1. 
     
     
         40 . The method of  claim 36 , wherein the antibody or antibody fragment has a ratio of binding affinity to the Ax1 protein at a pH of 5.8 to 7.0 of a tumor microenvironment to a binding affinity to the Ax1 protein at a pH of 7.2 to 7.6 in a non-tumor microenvironment of at least about 3:1.